I have already performed molecular identification 16 DNA on my actinobacteria isolate. However, I couldn't identify the species because the blasting result gave me 100 of similarity with more than 130 strain belonging to Streptomyces genus. How can I proceed to identify this Streptomyces bacteria ?
16S rRNA sequencing and 16S rDNA sequencing is one and the same.. We send our samples to First base, Malaysia and Scigenom Labs, India. The quality value of sequencing is good here.. We do 16S rDNA sequencing for molecular characterisation. DNA extraction we do manually or use kit.
You can read my research article if you need where I have described the manual procedure for DNA extraction from Actinomycetes.
Here is the link..
For identifiction of Actinomycetes , you have to isolate DNA and ensure it is not contaminate with proteins, RNA etc. Select the Primers for DNA, specifically for actinomycetes (Universal Primers are available from many laboratories). Go through the PCR amplification for DNA. After the DNA amplfication sent the samples for sequencing labs. You may get the DNA code sequences for actinomycetes. After that do BLAST analysis in Genbank for similarity search with available species. or you may simply send the isolated Actinomycetes DNA to sequencing labs for 16s RNA Seqencing.
Before molecular identification of Actinobacteria you can examine morphological and physiological tests such as temperature growth and pH, carbon and nitrogen consumption and spore chain morphology. 16S rRNA gene sequencing quality can be improved by purification of your samples before sending for sequencing. You can use gel extraction kit that enhances the quality of PCR sample. Also the company that you send your sample for sequencing is very important. South Korea usually presents high quality of sequencing
Thank you Dr Mona ,
I have already sent the The 16 Dna for sequencing in South Korea and I have blasted
in NCBI data base ,However , I couldn' t identify the species, ( 100 similarity with a lot of species belonging to streptomyces Groupe).
MY QUESTION IS Which specific phenotypic test should I do to identify this strain.
Best regards
The best course of action to take would be to read up articles where novel species of streptomyces have been characterised. These articles would give you an idea of what characterisation tests have been used previously for members of this genus.
I like to think of Actinobacteria as the fungi of bacteria, which is sort of resembled in the name. They can produce some interesting colony morphologies, in some cases filamentous like fungi and can also resemble fungi when viewed under a microscope with cells in long chain or filamentous formations sometimes also producing hyphal sporolating structures. However these features only appear in some Actinobacteria while others usually resemble bacteria from other phyla. With this in mind morphological characteristics both colony and cell would be important characteristics to analyse.
Other general characteristics of Actinobacter:
General stain gram positive
High GC content 60-70%
Usually mesophilic, non-thermophilic
soil or marine related
16S rRNA sequencing and 16S rDNA sequencing is one and the same.. We send our samples to First base, Malaysia and Scigenom Labs, India. The quality value of sequencing is good here.. We do 16S rDNA sequencing for molecular characterisation. DNA extraction we do manually or use kit.
You can read my research article if you need where I have described the manual procedure for DNA extraction from Actinomycetes.
Here is the link..
What kind of Molecular Identification technique can Identify actinomycetes isolates to species level?. Available from: https://www.researchgate.net/post/What_kind_of_Molecular_Identification_technique_can_Identify_actinomycetes_isolates_to_species_level [accessed May 26, 2017].
http://dx.doi.org/10.3389/fmicb.2016.00347/full
I suggest you to read the International Streptomyces Project and the
Guide to the classification and identification of the actinomycetes and their antibiotics. Here you have a key to Streptomyces species, maybe it can help you.
https://openlibrary.org/books/OL6137629M/Guide_to_the_classification_and_identification_of_the_actinomycetes_and_their_antibiotics
Good luck!!
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